Enzymatic aspects that modulate ion transport in cardiac muscle will be stressed with a major goal, mechanism of digitalis action in heart. The Na, K ions-ATPase will be studied as related to positive inotropic effect. The nature of H3-cardiac glycoside-enzyme interaction and dissociation will be also stressed. To accomplish this, isolation procedures will be modified to obtain a high specific activity and pure cardiac Na, K ions-ATPase. Comprehensive kinetic characteristics of the purified system will be carried out. These include: relationship of binding to inhibition; modulation of binding by ions; use of antibodies to intact Na, K ions-ATPase and components; studies to determine the conformational nature of digitalis-Na, K ions-ATPase interaction. The role of phosphoprotein metabolism (catalyzed by protein kinase) in Na, K ions-ATPase and modulation by Ca ions and ouabain will be studied. SAR studies of digitalis-enzyme interaction will be continued, stressing H3-glycoside binding. Modification of glycoside structure will be attempted so that a tightly bound (possibly covalent) digitalis-enzyme complex can be formed to be used in digitalis-site analysis. Reconstitution experiments employing protein and lipid components of the purified enzyme will include the use of artificial bilayer technique. The goal is to define components or sites with which glycosides interact. Purified Na, K ions-ATPase will be combined with SR to restore digitalis-sensitivity to a Ca ions exchange system. The possible role of the CAMP-dependent protein kinase will be studied in SR and in isolated purified cardiac troponin with respect to Ca ions modulation and digitalis action. Antibiotic ionophore, RO 2-2985/1 will be studied in detail in relation to clinical usefulness in heart failure and cardiogenic shock. A new, easily-reversible glycoside (AY 22-241) will be studied on Na, K ions-ATPase. The integrated roles of the Na, K ions-ATPase, SR and relaxing proteins should be of value in interpreting the diseased cardiac cell.